Genetic aberrations affecting Wnt signaling have been directly implicated in human cancer. Recent studies have identified frizzled (fz) proteins as Wnt receptors, but the complexity of ligand receptor interactions elucidated to date suggests that other components remain to be defined. Identification of the critical effectors of Wnt transforming action, as well as the specific consequences of Wnt activation will require further investigation. Aims of this grant include exploration of novel functions (uncovered by the applicant and his colleagues) of a prototype Wnt receptor, Hfz1, structure/function analysis of this and other fzs, and immunoaffinity purification and identification of proteins in the receptor complex. The applicant would then use this knowledge to investigate novel mechanisms of Wnt activation in tumor cells. A second major aim would be to elucidate mechanisms of transformation by Wnt and/or increased beta-catenin levels through investigation of alterations induced in normal cells. The role and contribution of Wnt target gene activation in the transformed phenotype in vitro and in vivo will be explored, and the applicant will also search for novel effectors by application of expression array analysis. The final aim will be to characterize new mechanisms of Wnt signaling activation in breast and other human tumor cells. This aim is supported by the applicant's identification of human tumor cell lines with elevated uncomplexed beta-catenin levels in the absence of genetic lesions affecting APC or beta-catenin. Wnt inhibitors, also characterized by the applicant, will be used to establish evidence of autocrine transforming or other novel mechanisms of Wnt activation. Finally, the effects of downregulation of Wnt signaling in human tumor cells could provide insights into the molecular basis for activation of the Wnt pathway in tumor evolution as well as the potential for therapeutic intervention.